Since human infection by avian influenza occurred in Hong Kong in 1997, outbreaks have been recently increasing in China and Southeast Asian countries such as Vietnam and Indonesia. Besides, at the time of co-infection with both human influenza and avian influenza, there has been high possibility of emergence of influenza virus variants which may infect people due to gene shift between the two influenza viruses (Chen et al., 2008). In recent years, it has been reported all over the world that the new H1N1 influenza virus derived from swine viruses infected and killed people, and the World Health Organization (WHO) also has warned of the outbreak of a new type of epidemic influenza virus and recommended each country to establish countermeasures against this possibility (WHO, 2009; Garten et al., 2009; Smith et al., 2009). There is still no effective preventive vaccine worldwide. Tamiflu, as the treatment that is currently in use, is in short supply and is produced at an enormous cost. Moreover, Tamiflu resistant viruses have been recently discovered, with the result that, when a new influenza pandemic arises, the technology for treating it is absent in the world.
In the present, reverse genetic pol I-pol II promoter systems used all around the world employ a human-derived promoter (Hoffmann et al., 2000). The thus developed human promoter systems are effectively operated in 293T cells, which are human-derived cell lines, to effectively create viruses, but the recombinant virus to be used for producing human vaccines cannot use cancer cells from human. Therefore, the Food and Drug Administration (FDA) in USA and the WHO have recommended Vero cells derived from African green monkeys (WHO, 2005). Although the human-derived pol I and pol II promoters operate the Vero cells derived from monkeys to some degree, the virus recovery rate is low as compared with for human-derived cells. Due to this low recovery rate, France tried to use chicken-derived pol I and pol II promoters to increase the virus recovery rate. Dr. Kawaoka's group in the USA succeeded in finding promoters derived from Madin-Darby Canine Kidney (MDCK) cells, and thus obtained research results of increasing the virus recovery rate (Massin et al., 2005; Murakami et al., 2008).
Hence, the present inventors endeavored to provide a reverse genetic method of increasing the recombinant virus recovery rate by using the pol I promoter derived from Vero cells, and as the result, they found the pol I promoter derived from Vero cells and confirmed that the virus recovery rate can be increased by using the vector comprising the Vero pol I promoter, and then completed the present invention.